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Cisplatin-induced expression of Gb3 enables verotoxin-1 treatment of cisplatin resistance in malignant pleural mesothelioma cells
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Clinical chemistry.
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Clinical chemistry.
Umeå University, Faculty of Medicine, Department of Medical Biosciences, Clinical chemistry.
Umeå University, Faculty of Medicine, Department of Odontology.
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2010 (English)In: British Journal of Cancer, ISSN 0007-0920, E-ISSN 1532-1827, Vol. 102, no 2, 383-391 p.Article in journal (Refereed) Published
Abstract [en]


A major problem with cisplatin treatment is the development of acquired-drug resistance of the tumour cells. Verotoxin-1 (VT-1) exerts its cytotoxicity by targeting the membrane glycolipid globotriasosylceramide (Gb3), a molecule associated with drug resistance. Cisplatin- and VT-1-induced apoptosis involves mitogen-activated protein kinase (MAPK) activation, and deactivation of MAPKs is associated with cisplatin resistance. This study aimed to investigate whether a sub-toxic concentration of VT-1 could enhance cisplatin-induced apoptosis and overcome acquired-cisplatin resistance in cultured cancer cell lines.


P31 and H1299 cells with corresponding cisplatin-resistant sub-lines (P31res/H1299res) were incubated with VT-1 and/or cisplatin followed by determination of Gb3 expression, cell viability, apoptosis, and signalling pathways.


Cells from the resistant sub-lines had elevated Gb3 expression compared with the parental cell lines, and cisplatin further increased Gb3 expression, whereas VT-1 reduced the percentage of Gb3-expressing cells. Combination of cisplatin and sub-toxic concentrations of VT-1 led to a super-additive increase of cytotoxicity and TUNEL staining, especially in the cisplatin-resistant sub-lines. Blockade of Gb3 synthesis by a Gb3 synthesis inhibitor not only led to eradicated TUNEL staining of P31 cells, but also sensitised P31res cells to the induction of apoptosis by cisplatin alone. Cisplatin- and VT-1-induced apoptosis involved the MAPK pathways with increased C-Jun N-terminal kinase and MAPK kinase-3 and -6 phosphorylation.


We show the presence of Gb3 in acquired-cisplatin resistance in P31res and H1299res cells. Cisplatin up-regulated Gb3 expression in all cells and thus sensitised the cells to VT-1-induced cytotoxicity. A strong super-additive effect of combined cisplatin and a sub-toxic concentration of VT-1 in cisplatin-resistant malignant pleural mesothelioma cells were observed, indicating a new potential clinical-treatment approach.

Place, publisher, year, edition, pages
Nature Publishing Group , 2010. Vol. 102, no 2, 383-391 p.
Keyword [en]
acquired resistance, apoptosis, cisplatin, Gb3, mesothelioma, verotoxin-1
URN: urn:nbn:se:umu:diva-3155DOI: 10.1038/sj.bjc.6605467ISI: 000273728500019OAI: diva2:141640
Available from: 2008-05-05 Created: 2008-05-05 Last updated: 2011-03-21Bibliographically approved
In thesis
1. Bacterial toxins for cancer treatment
Open this publication in new window or tab >>Bacterial toxins for cancer treatment
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Even though anti‐cancer chemotherapy has been continuously improved during the last decades. problems with adverse effects and drug resistance still constitutes a considerable obstacle and sets a demand for new effective treatment options. Tissue homeostasis in multi‐cellular organisms is maintained through intrinsic cell death, apoptosis, which removes unwanted or damaged cells. Disrupted apoptosis is an important factor in tumorgenesis and drug resistance, therefore induction or restoration of apoptotic pathways is also important for the treatment of cancer. Several naturally occurring bacterial toxins have the ability to induce apoptosis and could thus be candidates to complement or improve the therapeutic effect of other anticancer drugs.

The bacterial toxins, adenylate cyclase (AC) toxin from Bordetella pertussis, α‐toxin from Staphylococcus aureus and verotoxin‐1 (VT‐1) from Escherichia coli were investigated for their ability to induce apoptosis in different tumor cell lines. Toxin induction of cell death was investigated by cell viability assays, end‐stage apoptosis induction by DNA‐fregmentation (TUNEL) assay. Toxin receptor expression and signal transduction pathways to apoptosis were investigated by flow cytometry, caspase enzyme activity assays and western blot. Immunohistochemistry was used for identification of toxin receptor expression in tumor tissue samples.

AC‐toxin was cytotoxic and induced apoptosis in cultured malignant plural mesothelioma (MPM) and small‐cell lung cancer (SCLC) cells. Low‐toxic concentrations of AC‐toxin enhanced cisplatin cytotoxicity and apoptosis in both cell lines.

MPM‐cells with acquired cisplatin resistance were more sensitive to α‐toxin than the less resistant parental MPM cell line. A low‐toxic concentration of α‐toxin re‐sensitized resistant MPM cells to cisplatin cytotoxicity by apoptosis induced through the mitochondrial pathway without detectable activation of common up‐stream apoptosis signalling proteins.

VT‐1 was highly cytotoxic and induced apoptosis in globotriosylceramide (Gb3) ‐expressing glioma, breast cancer and non‐small‐cell lung cancer (NSCLC) cells but was not cytotoxic to non‐Gb3‐expressing cells. PPMP, an inhibitor of glucosylceramide synthesis which makes exposed cells unable to synthesize Gb3 rendered Gb3‐expressing cells resistant to VT‐1. MPM cells with acquired‐cisplatin resistance expressed Gb3 in contrast to the absent of expression in the less resistant parental cell line. Gb3, could however be up‐regulated by cisplatin in Gb3‐negative MPM‐cells. Presence of a low‐toxic concentration of VT‐1 potentiated cisplatin‐induced cytotoxicity and apoptosis in the cisplatin‐resistance MPM cell line. VT‐1 was a potent inducer of apoptosis, probably via stress‐induced Mitogen‐activated protein kinase (MAPK)‐signaling involving c‐Jun N‐terminal kinase (JNK) and p38, leading to disruption of the mitochondrial membrane integrety, activation of caspase‐9 and ‐3, and ultimately DNA fragmentation and cell death. Gb3 expression was demonstrated in clinical specimens of glioblastoma and breast cancer making these tumor types interesting for further VT‐1 studies.

We conclude that bacterial toxins may be used to induce apoptosis in several types of cancer cells. Low concentrations of verotoxin‐1 and α‐toxin may potentially be used to overcome acquired cisplatin‐resistance in cancer patients.

Place, publisher, year, edition, pages
Umeå: Medicinsk biovetenskap, 2008. 47 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1178
Alpha‐toxin, AC‐toxin, mesothelioma, lung cancer, glioma, breast cancer, caspases, MAP-Kinase, verotoxin‐1, cisplatin, apoptosis, Gb3, drug resistance
National Category
Other Clinical Medicine
urn:nbn:se:umu:diva-1637 (URN)978-91-7264-566-0 (ISBN)
Public defence
2008-05-23, Sal D, 1D, 9 trappor, Norrlands universitetssjukhus, Umeå, 13:00 (English)
Available from: 2008-05-05 Created: 2008-05-05 Last updated: 2010-01-21Bibliographically approved

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Johansson, DavidAndersson, CarolaMoharer, JasminJohansson, AndersBehnam-Motlagh, Parviz
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